The assimilation, storage and disposition of nutrient energy constitute a complex homeostatic system central to the survival of metazoa. In vertebrates, and particularly among land dwelling mammalian species, the ability to store large quantities of chemical energy in the form of adipose tissue triglycerides is crucial for surviving long periods of food deprivation. In order to maintain such food stores without sustaining continual alterations in the size and shape of the organism, a balance between energy intake and expenditure must be achieved. Despite intensive investigation, the molecular mechanisms which regulate energy intake and energy expenditure remain to be elucidated. It is anticipated that the identification of molecules that transduce nutritional information and regulate these functions will be critical to our understanding of the regulation of body weight in health as well as in disease states such as cancer. It is the objective of this application to isolate two of the genes which regulate this system: the mutant mouse genes obese (ob) and diabetes (db). These recessive mutations cause a severe obese phenotype which resembles human obesity in a number of critical aspects. The genes will be cloned using the techniques of positional cloning. RFLPs which flank these mutations are currently being used to initiate chromosome walks using yeast artificial chromosomes to isolate the ob and db genes. Candidate genes which have been genetically localized to the regions of ob and db will be tested for their ability to complement the mutations when introduced into transgenic or ES cell-derived mice. Once the genes have been isolated, a series of experiments will be initiated to determine the mechanism(s) whereby these genes control body weight. These studies will include a computer analysis of the predicted protein sequence, analysis of the sites of synthesis of the encoded RNA and protein, isolation of the human homologues of ob and db analysis of the regulation of the genes under physiologic and pathophysiologic circumstances, and functional studies of the encoded proteins in vitro and in vivo.